Myasthenia gravis (MG) is an autoimmune disease, caused by auto-antibodies to proteins in the postsynaptic membrane at neuromuscular synapses. Most MG patients carry antibodies to acetylcholine receptors (AChRs), the neurotransmitter receptor at vertebrate neuromuscular synapses (1,2,3). Auto-antibodies to AChRs are largely of the IgG1 and IgG3 subclass (4), which cause muscle weakness by three mechanisms: complement-mediated membrane lysis (5); crosslinking and depletion of cell surface AChRs (6); and to a lesser extent, functional blocking of the ACh-binding site (7). The ability of antibodies to AChRs to recruit complement, dimerize and modulate AChR expression is an important component of their pathogenic mechanism, since animals with experimental autoimmune MG (EAMG) can be rescued from disease with monovalent Fab fragments generated from AChR IgG antibodies and complement-deficient mice are protected against EAMG (8,6,9).
Approximately 20% of patients with MG lack antibodies to AChRs, and approximately 40% of these AChR-negative patients carry auto-antibodies to Muscle specific kinase (MuSK), a receptor tyrosine kinase that is essential for all aspects of synaptic differentiation and maintenance (10,11,12). The synaptic defects in MuSK MG overlap with those in AChR MG, including a reduction in the number of functional AChRs at synapses and unreliable synaptic transmission, resulting in muscle fatigue and weakness. In contrast to AChR MG, MuSK MG is caused in large part by IgG4 antibodies (13,14,15) that fail to engage complement and are considered functionally monovalent (13,14,15,16). Consequently, the accumulation of complement and muscle membrane damage, hallmark pathological features of AChR MG, appear insignificant in MuSK MG (13,17). Despite the paucity or absence of complement and cell damage in MuSK MG, the structural and functional deficits of synapses are extensive in MuSK MG, which highlights the key role that MuSK plays in organizing all aspects of synaptic differentiation (10,18).
AChR clustering and synapse formation are orchestrated by neuronally released Agrin, which binds to Lrp4, a member of the low density lipoprotein-related protein family, causing Lrp4 to bind and activate MuSK (19,20,21). Once tyrosine-phosphorylated, MuSK recruits Dok-7, an adaptor protein that becomes phosphorylated and recruits additional signaling molecules essential for synapse formation (22,23,24).
The extracellular region of MuSK contains three Ig-like domains and a Frizzled-like domain (10). The first Ig-like domain in MuSK is required for MuSK to bind Lrp4. Mutation of a single residue, I96, on a solvent exposed surface of the first Ig-like domain, prevents MuSK from binding Lrp4 and responding to Agrin (25,21). A hydrophobic surface on the opposite side of the first Ig-like domain mediates MuSK homodimerization, essential for MuSK trans-phosphorylation (25). Although MuSK is expressed by muscle and not by motor neurons, MuSK is essential for presynaptic as well as postsynaptic differentiation (10). In mice lacking MuSK, motor axons fail to stop and differentiate and instead wander aimlessly throughout the muscle (11). MuSK regulates presynaptic differentiation by clustering Lrp4 in muscle, which functions bi-directionally by serving not only as a receptor for Agrin and a ligand for MuSK, but also as a direct retrograde signal for presynaptic differentiation (26). In addition to its role during synapse formation, MuSK is also required to maintain adult synapses, as inhibition of MuSK expression in adult muscle leads to profound defects in presynaptic and postsynaptic differentiation (27,28).
In view of the above, a better understanding of MuSK activity and its role in the neuromuscular synapse in healthy and pathological states is an objective that serves a profound need. The need is underscored by the fact that MuSK MG is a debilitating autoimmune disease and one third of MuSK MG patients experience a life-threatening respiratory crisis. Long-term immunosuppression is the only current treatment option and the side effects of such an approach are considerable.
The citation of references herein shall not be construed as an admission that such is prior art to the present invention.